The bacterial induction of homograft sensitivity. 3. Effects of group A streptococcal membrane antisera.

Immunization of rabbits with Group A Type 12 streptococcal cell membranes has elicited serum antibodies which have the ability to cause rapid rejection of skin allografts in guinea pigs. Intradermal injection of such antisera has resulted in skin reactions characterized by prominent polymorphonuclear leukocyte infiltrates, similar to those noted in the Arthus reaction. The combined use of membrane antisera and epinephrine has resulted in hemorrhagic necrosis of the skin of guinea pigs. The ability of membrane antisera to exert these effects appears to be dependent upon the presence in the host tissues of antigen(s) shared by or cross-reacting with streptococcal membrane antigens. Such cross-reacting antigens may have a group distribution in the outbred guinea pig population. The results highlight the potential biological importance of antigens present in the Group A streptococcal membrane in the induction of altered tissue reactivity in the mammalian host.

Histocompatibility studies in a closely bred colony of dogs. 3. Genetic definition of the DL-A system of canine histocompatibility, with particular reference to the comparative immunogenicity of the major transplantable organs.

The segregation of the canine DL-A leukocyte group antigen(s) b, c, d, e, f, g, h, k, l, and m has been traced in 141 consecutive matings in the Cooperstown Colony of beagles. All of the leukocyte antigen(s) were regularly transmitted en bloc from parent to offspring, with no instance of independent segregation. A total of 23 haplotypes, including six different DL-A antigen patterns (gl, bkhfm, bkcd, e, be, fgl) was observed. 31 different DL-A phenotypes were observed in a population of 100 mongrel dogs. A number of statistically significant positive and negative associations between individual DL-A antigenic components occurred in this population. The results support the concept of the DL-A system as a complex immunogenetic system governed by a single region (or locus) of an autosomal pair of chromosomes. Studies of skin, kidney, heart, and liver allografts in the Cooperstown Colony indicated that the longest allograft survivals occur under genetically and serologically defined conditions of donor-recipient DL-A compatibility. Skin and renal allografts generally behaved in parallel fashion, while cardiac allografts survived for longer periods of time (MST = 47.1 days) than kidneys (MST = 28.1 days) or skin (MST = 25.1 days) under conditions of DL-A identity. Heart transplants were rejected at a more rapid rate than kidney, however, in DL-A-incompatible donor-recipient combinations. Liver transplants were accorded the longest survival time (MST = 76.2 days) under conditions of DL-A identity, but were rejected at a rapid rate (MST = 5 days) in DL-A-incompatible nonlittermate donor-recipient pairs. The results provide further evidence that the DL-A system is the principal system of histocompatibility in the canine species. The differences in survival of different organs under similar conditions of donor-recipient DL-A compatibility suggest, however, the existence of a number of unknown variables which may also be capable of significantly affecting allograft behavior.

Immunological effects of thermal injury. I. Inhibition of spermatogenesis in guinea pigs.

PIP: In the present study, controlled burns of 1 testis have induced the formation of testis-specific antibodies in Hartley guinea pigs. The antibodies reacted with autologous as well as homologous testicular extracts. In addition, pathological changes have been noted in the germinal components of the contralateral testis, which were similar to those observed after the induction of experimental allergic orchitis by active immunization with testicular tissue. Results of the study indicate that thermal injury to guinea pig testes can induce an autoimmune response similar to that observed after immunization with autologous or homologous testicular tissue. The antibodies formed were organ-and species-specific against a testicular antigen. Thermal injury may be associated with autoimmunization of the host by the injured organ.^ieng

Histocompatibility studies in a closely bred colony of dogs. I. Influence of leukocyte group antigens upon renal allograft survival in the unmodified host.

The establishment of a closely bred colony of beagles with known leukocyte group phenotypes has permitted an assessment of the role of leukocyte group antigens in conditioning the survival of renal allografts in the unmodified host. 22 kidney transplants obtained from leukocyte group-compatible donors were accorded a mean survival time of 25.5 days, as compared with 13.1 days for 27 transplants obtained from incompatible donors. Donor-recipient coefficients of correlation and Swisher erythrocyte group incompatibilities did not appear to affect the observed results. The mean survival time of 21 renal allografts performed in randomly selected mongrel dogs was 9.5 days. Availability of a carefully characterized and phenotyped canine population may be useful in further studies of the comparative immunogenicity of the major transplantable organs, and of methods designed to facilitate prolonged organ transplant survival in the mammalian host.

Histocompatibility studies in a closely bred colony of dogs.

17 Cooperstown beagles of known DL-A genotypes were exposed to supralethal total body irradiation and received a bone marrow allograft from a DL-A-identical donor; 11 littermate and 6 nonlittermate donor-recipient pairs were studied. The recipients are surviving uneventfully for 315, 364, 424, 440, 531, 531, 584, 605, 625, 635, and 649 days in the littermate group and for 211, 279, 280, 368, 479, and 480 days in the nonlittermate group. The radiation chimeras underwent bilateral nephrectomy and received a kidney allograft obtained from their respective marrow donor within 43-120 days after bone marrow transplantation. The renal allografts are surviving for 191, 200, 221, 234, 313, 349, 361, 377, 378, 405, 441, 444, 482, 557, 580, 581, and 586 days, respectively. 12 of 13 skin allografts obtained from the marrow donor are at present surviving in the recipients for 107, 110, 110, 110, 116, 122, 128, 143, 143, 162, 178, and 199 days, respectively; one graft was rejected at 84 days. In contrast, the radiation chimeras rejected 25 skin allografts obtained from DL-A-incompatible donors within 10.5-21 days (MST = 15.2 days). Skin transplants obtained from DL-A-identical siblings of the bone marrow donors were rejected within 20-36 days (MST = 25.8 days) in recipients of bone marrow cells obtained from littermate donors. Recipients of nonlittermate bone marrow transplants accorded such allografts a prolonged survival time of 27-76 days (MST = 56.2 days). Prospective selection of genotypically DL-A-identical donor-recipient pairs results in the regularly reproducible long-term survival of bone marrow allografts. The radiation chimeras produced in this manner have developed a donor-specific state of unresponsiveness to kidney and skin allografts. The results are consistent with the existence in the canine species of at least three closely linked genetic systems relevant to transplantation, including DL-A, MLC, and a possible bone marrow transplantation locus.

Histocompatibility studies in a closely bred colony of dogs. V. Mechanisms of cellular adaptation in long-term DL-A identical radiation chimeras.

20 Cooperstown beagles of known DL-A genotypes (B" dogs) were exposed to supralethal total body irradiation and received a bone marrow allograft from a DL-A identical donor (A" dog); the resulting chimeras have survived uneventfully for 882, 1466 days, with no evidence of secondary disease, and have been tolerant to kidney and skin allografts obtained from the donor of marrow. Treatment of "A" dogs with serum obtained from their long-term "B" chimeras had no significant effect upon the ability of the recipients to reject "B" skin allografts...

Ranks of donor-recipient histocompatibility for human transplantation.

Correlation of leukocyte groups with skin and renal allograft survival indicates that ranks of histocompatibility based upon current genetic concepts of the HL-A system may provide an approach to the selection of optimally compatible subjects for clinical organ transplantation. Such ranks may be expressed as a net histocompatibility ratio (NHR) between prospective donors and recipients. The best clinical results have been when this ratio is of 0.5 to 1. Donor-recipient compatibility situations where the ratio was 0.25 or less have been associated with a high incidence of transplant failure, regardless of whether the organ source was a living, related donor or a cadaver donor.

Heterophile antibodies in human transplantation.

Sensitization of human recipients with transplantation antigens (leucocytes, skin, or kidney allografts) has resulted in the appearance of serum hemagglutinins directed against sheep, guinea pig, and rat erythrocytes. Such hemagglutinins have been identified as IgG and IgM antibodies. Their appearance was not related to AB0 erythrocyte group incompatibility between donors and recipients, and the antibodies were not of the Forssman or Paul-Bunnel type. The antibody responses appeared to be primarily directed against antigen(s) present on rat erythrocytes, but shared to varying extents by other species. The peak antibody titers occurred in association with allograft rejection. In this regard, they may be of interest as a possible early warning system for the diagnosis and prompt management of rejection crises in clinical organ transplantation.

Erythrocytes in human transplantation: effects of pretreatment with ABO group-specific antigens.

Erythrocyte group antigens A and B can act as potent and group-specific transplantation antigens in man. ABO group-incompatible recipients pretreated with such antigens have rejected skin allografts obtained from donors incompatible for the same antigens in an accelerated (4-5 days) or white graft manner. Skin grafts applied to the same recipients from ABO-compatible donors were accorded first-set survival times. Intact erythrocyte suspensions and antigens isolated from hog (A substance) and horse (B substance) stomachs, were equally capable of inducing this type of allograft sensitivity. The latter observation broadens the spectrum of heterologous antigens capable of inducing allograft sensitivity in the mammalian host and provides a readily available, heat-stable, and water-soluble source of antigens for further studies of allograft rejection mechanisms in man.

Role of Ia-like products of the main histocompatibility complex in conditioning skin allograft survival in man.

This report correlates the survival time of 93 intrafamilial skin allografts performed under conditions of main histocompatibility complex (HLA) haploidentity with donor-recipient compatibility for products of the HLA-A, -B, -C, and -DR, as well as C3 proactivator, Glyoxalase I, and P loci located on the human 6th chromosome. Incompatibilities for HLA-A and -B (and to a lesser extent for HLA-C) and(or) for HLA-DR products exerted a strong influence upon the fate of skin allografts. When HLA-A and -B were considered alone, the most compatible group of grafts had a mean survival time of 15.8 d, as compared with 11.3 d for the most incompatible transplants. HLA-DR compatibility alone was associated with a mean survival time of 15.3 d, whereas HLA-DR-incompatible grafts had a mean survival time of 11.5 d. Incompatibilities for C3 proactivator, Glyoxalase I, and P did not have a significant effect upon graft survival. There was no evidence of an association between donor-recipient incompatibility at HLA-A, -B, or -C or at HLA-DR; such incompatibilities occurred independently of each other, in spite of the state of linkage disequilibrium known to exist between HLA-B and -DR. Incompatibilities for HLA-A, -B, and for HLA-DR exerted a potent additive effect upon graft survival. Skin grafts bearing one, two, or three incompatibilities had a mean survival time of 16.2, 13.7, and 10.7 d, respectively (P <0.0005).The results point to the important role played by the Ia-like products of the HLA complex (HLA-DR) in conditioning skin allograft survival in man. This consideration may be of direct relevance to the potential clinical usefulness of in vitro serological techniques for the detection of donor-recipient compatibility for HLA-DR.

Long-term study of vascularized free-draining intraperitoneal pancreatic segmental allografts in beagle dogs.

The purpose of the present study was to evaluate the significance of immunogenetic factors on the survival of pancreatic allografts in beagle dogs. Donors and recipients were leukocyte antigen (DLA)-typed and mixed lymphocyte culture (MLC)-tested. Recipients were made diabetic by total pancreatectomy and immediately implanted intraperitoneally with a vascularized, free-draining (duct unligated) pancreatic segmental (FDPS) allograft. Two groups of dogs were studied. In group I consisting of donor-recipient littermates, recipients were immunosuppressed with prednisone and azathioprine (n = 16 dogs), or not immunosuppressed (n = 4). In group II, recipients were made specifically unresponsive by total body radiation, autologous marrow implantation, and kidney transplantation from DLA-MLC identical donors, 1 yr before FDPS transplantation from the corresponding original kidney donors. Survival of the FDPS grafts in group I was inversely related to pretransplant MLC reactivity, irrespective of DLA genotyped match between donor and recipient. Thus, immunosuppressed high MLC reactors (n = 8) rejected FDPS grafts between 7 and 14 d, whereas immunosuppressed low MLC reactors (n = 8) accepted grafts for 25 to 260+ days, and nonimmunosuppressed low MLC reactors (n = 4) accepted grafts for 9-55 d. Rejection (hyperglycemia) of FDPS grafts was sudden, permanent, and unpredictable despite weekly intravenous glucose tolerance tests with measurements of glucose disappearance rates and serum insulin responses. Nevertheless, serial in vitro cell-mediated lymphocytotoxicity (CML) assays revealed increases in CML before graft rejection in low MLC reactors, and decreases in both CML and MLC responses before graft rejection in high MLC reactors. FDPS graft survival was indefinite (>6 mo) in group II dogs, despite low-grade MLC reactivity (2:4 dogs) and CML responses (4:4 dogs). Biopsies of FDPS grafts at 6 mo in normoglycemic dogs showed disappearance of exocrine tissue and coalescence of islets in both groups I and II, but with less fibrosis in group I (immunosuppressed). These results indicate that (a) pancreatic islets in vascularized grafts (FDPS) may survive indefinitely in the presence of a good tissue match best predicted by MLC testing, (b) tissue specific histocompatibility factors appear to be common enough between kidney and pancreas to allow for long-term survival of both organs transplanted from the same donor, at least in appropriate recipients (group II), and (c) immunosuppression is associated with less fibrosis in FDPS allografts.

Induction of allogeneic unresponsiveness in adult dogs. Role of non-DLA histocompatibility variables in conditioning the outcome of bone marrow, kidney, and skin transplantation in radiation chimeras.

Exposure to supralethal total body irradiation and transplantation of bone marrow from a DLA- and pedigree-identical donor have regularly produced successful engraftment and the establishment of stable long-term chimerism in beagles of the Cooperstown colony. Bone marrow allografts performed in pairs of dogs bearing identical DLA haplotypes derived from different pedigree origins (i.e., different classes of the same haplotype) yielded two different results. Depending upon the particular haplotype pedigree combination used, such transplants either led to long-term chimerism or to failures of engraftment, secondary disease, and death of the recipients (i.e., pedigree-incompatible combinations). Radiation chimeras given bone marrow from a DLA-and pedigree-identical donor were challenged within 8-12 h after marrow transplantation with a renal allograft obtained from another DLA- and pedigree-identical donor. The recipients have remained unresponsive to such renal allografts and have survived indefinitely with normal renal function. In contrast, renal allografts obtained from donors bearing the same DLA haplotypes derived from pedigree-incompatible sources were rejected within 25-50 days after transplantation. The long-term surviving recipients have also been unresponsive to skin allografts obtained from their donor of marrow and the kidney donor. Skin grafts obtained from other DLA- and pedigree-identical dogs were rejected within 13-41 days, and grafts from DLA-incompatible donors survived for 10-25 days. These results highlight the potential importance of genetically controlled histocompatibility determinants other than DLA in conditioning allograft reactivity. The determinants uncovered in the present study appear to be linked to the DLA complex, as demonstrated by the ability of the pedigree origins of DLA haplotypes present in individual dogs to serve as an effective marker system for such non-DLA antigen(s). The results also point to the potential usefulness of the early postirradiation period for the induction of allogeneic unresponsiveness in large adult mammals.

The protective effect of calcium inhibitors and of captopril on the renal microcirculation during reperfusion.

Reperfusion injury is increasingly recognized as a key factor in the development of posttransplant acute tubular necrosis. Previous studies have shown that addition of the calmodulin inhibitor trifluoperazine (TFP) to Collins' flush solution protected the cortical microcirculatory integrity and dramatically improved renal viability after transplantation. The present report describes the protective effect(s) of TFP in the course of reperfusion injury. Twenty mongrel dogs underwent bilateral nephrectomy; in each instance, the left kidney was flushed immediately with 250 ml of cold Collins' solution, and the right kidney was flushed with the same solution containing TFP, 5 mg/L. After 48 and 72 hr of preservation, each kidney was connected through silastic shunts to the femoral vessels of another dog. The mean renal blood flow (RBF) immediately after reperfusion was 2.2 ml/g/min and 1.7 ml/g/min in the left and right kidneys, respectively, and was similar to mean RBF measurements prior to nephrectomy. After 15 min of reperfusion, there was a sharp decrease in mean RBF in the Collins' flushed kidneys, which persisted after 60 min of reperfusion (0.37 ml/g/min). In contrast, there was only a mild decrease in mean RBF in the TFP-flushed kidneys (1.27 ml/g/min). A partial explanation for the favorable effect of TFP may be related to the inability of the ischemic cell to handle the increased calcium load associated with reperfusion (calcium paradox). In a test of this possibility, 0.5 mg/kg of verapamil, a calcium channel blocker, was infused during reperfusion. No beneficial effects of this drug were noted in either Collins' or TFP-flushed kidneys (n = 10). However, when 1.25 mg/kg of captopril, an angiotensin-converting enzyme inhibitor, was infused at the time of reperfusion, a dramatic amelioration of the reperfusion injury occurred in the Collins' flushed kidneys (1.2 ml/g/min) (n = 10). Taken together, these data suggest that the damage to cold-preserved kidneys flushed with Collins' solution alone may occur at the time of actual reperfusion. Such reperfusion damage is ameliorated by TFP and captopril. The known relationship between calcium and the effect of angiotensin on the vascular smooth muscle cell may explain in part the protective role of calcium inhibitors placed in preserved kidneys prior to reperfusion.

Synergistic effects of combined immunosuppressive modulation. I. Unresponsiveness to dendritic cell-depleted renal allografts in dogs exposed to total-lymphoid irradiation.

Attenuation of the allogeneic stimulus provided by dendritic cells (DC) was achieved by irradiation of the donors, followed by their reconstitution with bone marrow from the prospective DLA-identical recipient. Following long-term (131-187 days) recovery free of graft-versus-host (GVH) disease, the chimeric kidneys were placed into the corresponding recipients; such allografts were rejected at 55, 55, and 60 days, respectively. Four other recipients were conditioned with 1750-1790 cgy of total lymphoid irradiation (TLI) and were then given a similar chimeric kidney from the corresponding partner. These allografts currently survive for 296, 295, 290, and 252 days, respectively. A third group of four dogs was exposed to TLI prior to transplantation of a normal DLA-identical kidney. These grafts were rejected at 20, 42, 46, and 242 days, respectively. Thirteen DLA-identical renal allografts transplanted into normal dogs survived for 13-38 days (mean survival time = 28.6 days). Depletion of allogeneic DC alone, or TLI alone, produced relative prolongations in allograft survival in canine recipients. Combined use of these two modalities, however, resulted in long-term allogeneic unresponsiveness in the recipients.

An approach to organ salvage from non-heartbeating cadaver donors under existing legal and ethical requirements for transplantation.

Effective utilization of nonheartbeating cadaver donor organs is limited by the time required to obtain the necessary family consent prior to organ retrieval (a delay of at least 4-6 hr); this exceeds by far the maximum tolerance of kidneys to warm ischemia. Measures that could theoretically permit use of such organs include: (1) rapid in situ flush cooling; (2) continued in situ kidney cooling until permission for donation is secured; and (3) cell-membrane stabilization of vital organs, with only minimal invasion of the donor body. These measures were tested experimentally in dogs. Hemorrhagic shock was produced in mongrel dogs. One hour after cessation of heartbeat, a rapid perfusion tube was placed into the femoral artery; it was advanced, and its balloon was inflated in the aorta above the renal vessels. The kidneys were then flushed in situ with 1000 cc of cold preservation solution containing a calmodulin inhibitor, trifluoperazine. Two other catheters were inserted percutaneously into the peritoneal cavity for continuous intraperitoneal cold perfusion. Core temperatures of 4 degrees C were maintained in situ in the kidneys for 5 hr. Six hours after cardiac arrest, the kidneys were removed and preserved ex vivo at 4 degrees C for 24 hr, and were then transplanted into their respective hosts (n = 11), where they sustained life uneventfully. This method requires a 2-inch incision in the groin of the prospective donor, and two small stab wounds of the abdomen; i.e., semi-invasive procedures which are commonly performed in emergency rooms. The perfused body could then be released to the family if donation is denied. The recently documented increased willingness of the public to donate organs when the termination of life support is not an issue, and court decisions that have authorized the performance of nondeforming diagnostic procedures in cadavers without consent, suggest that the salvage of transplantable semi-invasive procedures described in this study may be useful in helping to alleviate the current shortage of transplantable organs. This technique can provide the time needed for families to consider the option of organ donation from nonheartbeating cadaver donors in an unhurried and unpressured manner, while preserving the viability of vital organs during the decision-making process.

Effect of renal allograft dysfunction upon cyclosporine trough levels in host blood.

Cyclosporine (CsA) dose adjustment after renal transplantation is generally based on serum, plasma, or whole-blood trough level values. In the face of increased levels, the dosage is reduced in order to prevent CsA-induced nephrotoxicity. There is a paucity of data, however, on the kinetics of CsA in association with dysfunction of the transplanted kidney. This study documents dramatic rises in serum cyclosporine trough levels at the time of rejection crises, as well as following periods of nonimmunological allograft oliguria. Decreases in CsA dosage in such patients failed to result in a significant lowering in trough levels. Therapeutic CsA trough levels were generally at the 70-140 ng/ml level; at the time of rejection, the same doses of CsA resulted in a rise of trough levels to 300-500 ng/ml. As the rejection crises resolved and kidney function improved, the CsA serum trough levels returned to their lower levels. These results suggest that the urinary elimination of CsA and its metabolites may be a key determinant of CsA trough levels, and that the status of renal function at the time of testing must be considered in the interpretation of the data. In support of this observation, the CsA concentrations in 4-6 hr post-CsA-administration urine samples ranged from 400 ng/ml to 4500 ng/ml, as measured by high pressure liquid chromatography. The data suggest that rising CsA trough levels in a previously stable recipient may serve as a valuable early warning index of impending allograft dysfunction (rejection, infection, and obstruction). This appears particularly true during the first 30 days after renal transplantation, when the incidence of rejection is the greatest in this patient population.

Preoperative preparation of high-risk, specifically hyperimmunized canine renal allograft recipients with total-lymphoid irradiation and cyclosporine.

Hyperimmunized subjects are a particularly high-risk and rapidly growing group in the patient population awaiting renal transplantation. In a search for methods designed to ameliorate the prognosis in such cases, dogs of defined DLA genotype were sensitized with DLA incompatible skin allografts and injections of buffy coat. Each recipient was challenged with a renal allograft bearing the same DLA incompatibilities. Five dogs received kidney transplants, without any other treatment, and rejected their transplants at 2.5, 4, 5, 6, and 6.5 days, respectively. Another four dogs were given a 9-11-week course (1760 +/- 35 cGy) of total-lymphoid irradiation (TLI), followed by rabbit antithymocyte globulin (ATG); these animals rejected their renal allografts at 7, 8, 14, and 17 days, respectively. Five other dogs were treated with TLI and received cyclosporine (CsA) and methylprednisolone (MPd) daily until graft rejection. Their renal allografts survived for 7.5, 8.5, 20, 62, and 227 days, respectively. Renal allografts placed in normal recipients under the same conditions of donor-recipient DLA incompatibility had a mean survival time of 12.4 days (range: 10-18 days). At the time of transplantation, the specific anti-DLA antibody titers in the recipients were 81 to 243 in the untreated dogs; 27 to 81 in the TLI-ATG-treated group, and 3 to 243 in the TLI-CsA/MPd-treated group. The titers fell within 24-48 hr after renal transplantation, to 3 to 81 in the untreated sensitized dogs; they were 3 to 9 in the TLI-ATG-treated group, and were 9 to 243 in the TLI-CsA/MPd treated group. The cytotoxic antibody titers reached postoperative peaks of 6500 to 200,000 in the untreated dogs; 729 to 6500 in the TLI-ATG-treated dogs, and 243 to 6500 in the TLI-CsA/MPd-treated recipients. The combined use of TLI and CsA/MPd can significantly inhibit the capacity of immunized recipients to muster a secondary humoral response to the DLA antigen(s) used in the sensitization process; such treatment also abrogates the ability of the recipients to reject renal allografts bearing the same DLA specificities in accelerated fashion. This effect of TLI and cyclosporine may be of relevance to current severe problems in high-risk hyperimmunized human renal transplant candidates.

Immunohistologic analysis of human renal allograft dysfunction.

The value of percutaneous core needle biopsy in the immunohistological evaluation of renal allograft dysfunction was studied in 72 consecutive biopsies performed in 42 patients. The phenotypes of infiltrating cells mediating graft destruction were identified with monoclonal antibodies and immunoperoxidase staining techniques. Light microscopy, electron microscopy, and immunofluorescence staining were performed in all biopsies. Biopsies were divided into groups depending on their classification on the basis of standard histologic criteria, i.e., acute tubular necrosis (ATN), acute interstitial rejection, acute vascular rejection, chronic rejection and renal disease in native kidneys (RDNK) of nontransplant patients. Immunohistologic analysis of graft biopsies showed a significant increase in Leu 1 (pan-T cells), (P less than 0.001), Leu 2 (cytotoxic/suppressor cells) (P less than 0.001), and Leu 3 cells (P less than .05) in acute interstitial rejection. The expression of DR antigen was significantly increased in both acute (P less than .025) and chronic (P less than .05) rejection, when compared with the findings in ATN biopsies. Leu M1 (monocytes/activated T cells) and Leu 10 (B cells/macrophages) were significantly increased (P less than 0.05 and P less than .005, respectively) in acute interstitial rejection only. The helper/suppressor ratio of infiltrating cells showed no significant change in any clinopathologic category. There was no correlation between the cell populations infiltrating the graft and those monitored in the peripheral blood. Allograft mononuclear cell infiltrates in cyclosporine (CsA) vs. azathioprine-treated patients revealed significantly fewer Leu 2 (P less than .05) and Leu M1 (P less than .05) cell populations in CsA patients during acute rejection. In 32 of these 72 biopsies (44.4%), the biopsy results provided a direct contraindication to the use of steroids, by allowing differentiation between allograft rejection and other causes of graft dysfunction. A total of 38% of the biopsies yielded a histological diagnosis that contradicted the clinical pre-biopsy diagnosis. All allografts showing evidence of severe small vessel disease and/or antibody-mediated rejection eventually were lost. These data highlight the usefulness of needle biopsy material as a guide to the study of intragraft immune events and to clinical management of recipients.

Enhanced resistance to the effects of hypothermic ischemia in the preserved canine kidney.

Addition of trifluoperazine (TFP), a powerful calmodulin inhibitor to Collins' flush solution has exerted a significant protective effect on the cold-preserved kidney, with successful autotransplantation of 80% of preserved kidneys (4 of 5) after 72 hr of storage. In contrast, use of Collins' solution alone resulted in successful autotransplantation of only 33% (2 of 6) of kidneys after a similar period of preservation. In an attempt to analyze the significance of this result, the microcirculation of preserved kidneys was studied with injections of technetium-labeled microspheres into the kidneys, followed by study with a noninvasive radionuclide scintiphotography (RNS) technique that does not interfere with subsequent transplantation of the kidney. Such studies demonstrate that prolonged cold preservation after flush-cooling with Collins' solution is associated with a progressive deterioration of the integrity of the microcirculation, resulting in poor flow to the renal cortex. In contrast, when TFP is added to the Collins' solution, there are uniform increases in renal cortical flow in kidneys stored for 48 and 72 hr, with preservation of the integrity of the renal microcirculation. Biological testing shows a clear-cut correlation between these observations and the capacity of the tested kidneys to sustain life after retransplantation. Further experiments suggest that the decreases observed in cortical flow in kidneys preserved in the cold for 72 hr are a consequence of cellular swelling, and not of a vasospastic response. The data support the conclusion that TFP exerts its protective effect on the basis of its membrane stabilizing capacity as a calmodulin inhibitor, and not through direct vasodilatation.

Protective effects of trifluoperazine on the microcirculation of cold-stored livers.

Previous studies have shown a protective effect of trifluoperazine (TFP), a calmodulin inhibitor, upon the microcirculation of cold-stored kidneys. The present study points to similar beneficial effects of TFP on the microcirculation of cold-stored livers; 25 canine livers were preserved for 24 hr with Euro-Collins' solution (EC) (n = 8), University of Wisconsin solution (UW) (n = 7), or UW + TFP (n = 10). The stored livers underwent heterotopic transplantation (HLTX); hepatic-artery and portal-vein pressure and flow were monitored; oxygen consumption and extraction were measured before HLTX and at 15-min intervals after reperfusion, for 1 hr. Mean hepatic-artery and portal-vein flow (HAF & PVF) prior to donor hepatectomy were 172 and 530 cc/min, respectively. Poor HAF and PVF occurred in EC-HLTX (mean 35, 175 cc/min, respectively). The damaged EC-flushed livers could not compensate to the decreased hepatic blood flow by increased oxygen extraction (oxygen consumption and extraction, 8.7 vol.% and 48%, respectively). Light and electron microscopy showed severe liver necrosis and periportal hemorrhages. Improved hepatic-artery and portal-vein flows were seen in UW HLTX (105 and 254 cc/min), and oxygen consumption and extraction were 16.4 vol.% and 66%, respectively. Liver biopsy taken just before reperfusion revealed well-preserved liver architecture. Liver biopsy obtained 1 hr after reperfusion revealed marked edema of the portal triad, sinusoid congestion, and hemorrhage. Electron-microscopy biopsies obtained during reperfusion at 15-min intervals revealed severe vasospasm of the terminal hepatic arterioles and progressive damage to the liver microcirculation. The addition of TFP to the UW-flush solution resulted in excellent protection of the liver microcirculation. Marked increase in hepatic-artery and portal-vein blood flow was noted after reperfusion (mean 167 and 421 cc/min, respectively (P 0.02 vs. UW: P 0.001 vs. EC). The recovery of metabolic activity was evident by the high oxygen consumption and extraction (25.8 vol.% and 80%, respectively). And serial liver biopsies obtained after reperfusion have shown excellent protection of liver architecture and the absence of hepatic arteriolar vasospasm. Taken together, these data suggest that the addition of TFP to the UW solution protects the liver microcirculation by rendering the hepatic microcirculation insensitive to vasospastic stimuli during reperfusion, thus permitting better metabolic recovery after transplantation.